Data Overview

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that plays a vital role in biogeochemical cycling in coastal and marine environments. Of particular interest in our research is the flavin-containing monooxygenase (FMO) enzyme found in this organism. This specific sequence contains an N78S mutation, and a 6x His-tag for IMAC purification. This mutant Tmm is significant for our project because it possesses a Km of 7.3 μM which we believe makes it optimal for TMA sensing. This part has the wild-type start codon GGC (glycine)

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that plays a vital role in biogeochemical cycling in coastal and marine environments. Of particular interest in our research is the flavin-containing monooxygenase (FMO) enzyme found in this organism. This specific sequence contains an N78S mutation, and a 6x His-tag for IMAC purification. This mutant Tmm is significant for our project because it possesses a Km of 7.3 μM which we believe makes it optimal for TMA sensing. This part has ATG (methionine) inserted upstream of the wild-type starting coding GGC (Glycine) to experiment with which has the most optimal expression.

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that plays a vital role in biogeochemical cycling in coastal and marine environments. Of particular interest in our research is the flavin-containing monooxygenase (FMO) enzyme found in this organism. This specific sequence contains an N78S mutation, and a 6x His-tag for IMAC purification. This mutant Tmm is significant for our project because it possesses a Km of 7.3 μM which we believe makes it optimal for TMA sensing.This part has the wild-type starting coding GGC (glycine) replaced with ATG (Methionine) to experiment with which has the most optimal expression.

Roseovarius sp. 217 is a marine bacterium isolated from a methyl halide oxidizing enrichment culture derived from surface seawater near Plymouth, England. This species serves as a foundational component for my trimethylamine monooxygenase (TMM) research, as it has the capability to convert trimethylamine (TMA) into trimethylamine N-oxide (TMAO), which is then further processed into formaldehyde and ammonia. The sequence was modified with a 6x His-tag for IMAC purification.

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that play a vital role in biogeochemical cycling by converting TMA into TMAO. To synthesize M. aminisulfidivorans TMM for our project, the protein-coding sequence was inserted into a biological device. This construct utilizes a T7 promoter to initiate transcription, coupled with a ribosome binding site (RBS) that facilitates protein synthesis and interacts with the enzyme-coding sequence itself. This variant retains the wild-type start codon GGC (glycine). Gene expression is regulated and properly stopped using a T7 terminator. The 6x His-tag will be used to find the protein using IMAC.

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that play a vital role in biogeochemical cycling by converting TMA into TMAO. To synthesize M. aminisulfidivorans TMM for our project, the protein-coding sequence was integrated into a biological device. This construct features a T7 promoter to initiate transcription, along with an ATG (methionine) codon inserted upstream of the wild-type GGC (glycine) start codon. This modification aims to provide a more recognizable start for translation. The ribosome binding site (RBS) promotes efficient protein synthesis while interacting with the enzyme-coding sequence. A T7 terminator is included to ensure proper transcription termination. The 6x His-tag will be used to find the protein using IMAC.

Methylophaga aminisulfidivorans are aerobic bacteria isolated from seawater near Mokpo, South Korea, that play a vital role in biogeochemical cycling by converting TMA into TMAO. To synthesize M. aminisulfidivorans TMM for our project, the protein-coding sequence was incorporated into a biological device. This construct employs a T7 promoter for transcription initiation, with the wild-type GGC (glycine) start codon replaced by ATG (methionine) to explore the effects of a more common start codon arrangement on expression. The ribosome binding site (RBS) enhances protein synthesis while interacting with the enzyme-coding sequence. A T7 terminator is used to terminate gene expression appropriately. The 6x His-tag will be used to find the protein using IMAC.

Roseovarius sp. 217, isolated from a methyl halide oxidizing enrichment culture in seawater near Plymouth, England, plays a critical role in biogeochemical cycling by converting TMA into TMAO. To synthesize the TMM enzyme from Roseovarius sp. 217, which contains the wild-type start codon GGC (glycine), the gene was incorporated into a biological device. This construct combines a T7 promoter to initiate transcription with a ribosome entry site to promote the initiation of protein synthesis and interaction with the enzyme coding sequence. Transcription termination is ensured with a T7 terminator. The 6x His-tag will be used to find the protein using IMAC.

Roseovarius sp. 217, isolated from a methyl halide oxidizing enrichment culture in seawater near Plymouth, England, plays a critical role in biogeochemical cycling by converting TMA into TMAO. To synthesize the TMM enzyme, a variant was created where an ATG (methionine) codon was inserted upstream of the wild-type GGC (glycine) codon. This gene was integrated into a biological device combining a T7 promoter, which initiates transcription, with a ribosome entry site that facilitates the start of protein synthesis. A T7 terminator ensures proper transcription termination. For purification, reverse purification will be employed, where everything except the target His-tagged molecule will be removed, allowing for the efficient isolation of the TMM enzyme.